Prediction of LC retention of steroids using solvatochromic parameters

Summary The proportion of organic modifier and pH of the hydroorganic mobile phase were optimized to separate a series of nine important steroids by LC. It has been shown that linear plots of logk′ against Reichardt’sE _T ^N parameter for the mobile phase is observed over the whole range of compositions of practical importance. Data from these plots summarize an approach for optimization of resolution from only two retention measurements for each compound. pH is shown to be irrelevant. This behaviour could be extrapolated to other steroids and drastically reduce the number of retention values needed for mobile phase optimization for screening analysis of steroids in doping investigations.     

反相液相色谱中溶质保留与溶剂化结构参数间的定量关系

本文研究了等度条件下反相液相色谱中溶质保留与溶剂化结构参数间的定量关系;考察了反相液相色谱保留值变化规律式lnk'=a+cC~b中参数a.c 与溶剂化结构参数间的定量关系,结果表明能较好地预测等度和不同冲洗剂组成下溶质的保留值,将乙腈-水和甲醇-水冲洗剂下的a.c值定量关联,表明在考虑溶质的氢键作用参数后,其相关性有所提高.     

Probing solute clustering in supercritical solutions using solvatochromic parameters

Kamlet and Taft polarity parameters are presented for three supercritical solutions containing solutes of different polarities at two concentrations. It is shown that the polarizability and hydrogen-bond-donating ability change significantly with pressure. In the lower pressure regime, naphthalene and salicylic acid cause a decrease in the polarity parameters compared with the pure fluid, whereas toluic acid causes an increase. These differences can be explained in terms of solute-solute and solute-indicator clustering. It is shown that there is, however, a direct correlation between the change in the hydrogen-bond-donating ability and polarizability for all solutes and all pressures and concentrations, which is thought to result from changes in the amount of solvent required to solvate the solute.     

Determination of polarity parameters for liquid crystals using solvatochromic method in anisotropic and isotropic phases

The use of liquid crystals (LCs) as anisotropic solvents is desired for various potential applications and usually for other organic and inorganic compounds. In this work, solvent polarity parameters are obtained using a spectroscopic method for four LCs with a range of high and low dielectric anisotropy (?ε). Solvatochromic polarity parameters for these LCs were defined via Kamlet–Abboud–Taft polarity functions characterizing different temperatures and phases, isotropic and anisotropic, and using the Reichardt’s dye and 2,6-diphenyl-4-(2,4,6-triphenyl-1-pyridinio) phenolate standard probe. The investigated polarity parameters reveal the effects of LC media on the photo-physical behaviour of solute molecules in isotropic and anisotropic media. Subsequently, a new LC polarity parameter (Z_o) is introduced as an overall matrix anisotropy polarity parameter to characterize variation between isotropic and anisotropic phases. The values of Z_o are sorted from higher to lower dielectric anisotropies (?ε).     

Prediction of retention data when using incremental multiple development techniques

Summary The apparentRF value of a compound after successive development steps in incremental multiple development (IMD) is determined. The model developed enables prediction of the migration and the zone width of investigated compounds in IMD with linearly and quadratically increasing development distances. For the experimental investigations we examined naturally occurring compounds as model substances on different stationary phases. The coincidence of predicted and measured chromatographic data strongly support our formulas. Presented at the 21^st ISC held in Stuttgart, Germany, 15^th–20^th September, 1996     

Trace determination of steroids causing age-related diseases using LC/MS combined with detection-oriented derivatization

With the rapid shift to an aging society in Japan, age-related diseases, such as osteoporosis, dementia and cancer, are sharply increasing. The measurement of steroids related to these diseases in biological fluids and tissues is useful for elucidation of the nature, diagnosis and treatment of such diseases. LC/MS is considered to be the most promising method for this purpose due to its specificity and versatility, but it sometimes does not demonstrate the required sensitivity for trace amounts of steroids, because steroids have a rather low response using either electrospray ionization (ESI) or atmospheric pressure chemical ionization (APCI). To overcome this problem, the author developed detection-oriented derivatization procedures for steroids in LC/MS. For ESI-MS, introduction of a permanently charged moiety is effective. Based on this, 2-hydrazino-1-methylpyridine was developed and used in monitoring prostatic 5alpha-dihydrotestosterone, a good index for the follow-up of patients affected by prostate cancer under androgen deprivation therapy and salivary dehydroepiandrosterone, which is now often designated as an anti-aging hormone. A proton-affinitive Cookson-type reagent, 4-[2-(6,7-dimethoxy-4-methyl-3-oxo-3,4-dihydroquinoxalyl)ethyl]-1,2,4-triazoline-3,5-dione, was used for the determination of 1alpha-hydroxyvitamin D3 [1alpha(OH)D3], a synthetic prodrug of the active form of vitamin D3, in human plasma, and this new LC/positive-APCI-MS method enabled the pharmacokinetic study of 1alpha(OH)D3 in humans. Electron-capture APCI-MS based on derivatization with 2-nitro-4-trifluoromethylphenylhydrazine was used for the analysis of neurosteroids, which affect brain excitability through action at the neurotransmitter receptors. With this method, the stress-induced rapid biosynthesis of pregnane-type neurosteroids in rat brains was demonstrated.     

Quantitative screening for steroids in animal feeding water using reversed phase LC with gradient elution

Several isocratic separations for the determination of 20 steroids (STER) in animal feeding water samples (AFWS) from drinking-trough by LC using a mobile phase ACN/H(2)O (35:65 v/v) and different RP columns (Hypersil C18, Gemini C18 (GM), Purospher Star C18, Synergi Max C12, and Synergi Fusion) and UV detection were obtained. The elution order was the same: a first group of corticoids (CC) was early eluted, a second group of CC and anabolics (AAS) exhibited intermediate retention, and a third group constituted by AAS was strongly retained. To improve the separation performances of the isocratic separations an ACN gradient elution optimization was carried out for each column. The most satisfactory results were obtained using a Purospher column which allowed the separation of 19 STER in an analysis time close to 26 min. After sample preparation using SPE, method validation was performed in an AFWS spiked with STER according to the EC decision criteria established for quantitative screening methods. For this purpose calibration graphs, extraction efficiencies, decision limits, detection capabilities, precision (repeatability and within-laboratory reproducibility), accuracy, selectivity, and robustness were evaluated. The proposed method was applied to other AFWS with satisfactory results.     

Sample preparation for the determination of steroids (corticoids and anabolics) in feed using LC

An improved sample preparation procedure for the determination of 17 steroids (corticoids (CC) and androgenic anabolic steroids (AAS)), used potentially as growth promoters, in feed samples has been developed. This procedure is based on two reported LC-UV methods. The improved procedure includes a leaching process using ACN, saponification, and SPE using polymeric cartridges. The proposed method was validated according to the EU criteria established for quantitative screening methods in PFS. The extraction efficiencies, decision limits (CCalpha) and detection capabilities (CCbeta), for these compounds were in the ranges of 82-100%, 19-40, and 24-53 microg/kg, respectively. The repeatability and the within-laboratory reproducibility at 1.0, 1.5, and 2.0 CCbeta levels were smaller than 10%. Accuracy was in the 97-101% range. The robustness was evaluated using the Youden robustness test. This method was applied to the analysis of steroids in different kinds of FS with satisfactory results.     

Kamlet-Taft solvatochromic parameters of aqueous binary mixtures oftert-butyl alcohol and ethyleneglycol

The , , and * Kamlet-Taft solvatochromic parameters have been determined for aqueous mixtures of tert-butanol and ethyleneglycol from spectroscopic data.     

Targeted identification of phosphorylated peptides by off-line HPLC-MALDI-MS/MS using LC retention time prediction

Protein phosphorylation is a type of posttranslational modification which plays an important role in cell regulation and signal transduction. Because of its biological relevance, a considerable amount of interest has been paid to the development of efficient techniques for phosphopeptide analysis. Although advances in MS control have enabled the high-throughput discovery of proteins from limited amounts of sample, automated selection of MS/MS precursor ions based on intensity alone can significantly hamper the detection of low-abundance phosphopeptides. On the basis of the observation that the introduction of a phosphate moiety does not dramatically change peptide retention time in reverse-phase chromatography, phosphopeptide specific MS/MS fragmentation attempts based on LC retention time and m/z were evaluated using a standard protein mixture, then using in vitro phosphorylated myelin basic protein. Results indicated that the majority (98%) of phosphopeptides identified eluted within a +/- 4-min window of the predicted LC elution time. While studies presented here are primarily proof of concept in nature, data suggest that the use of LC retention time prediction could be a valuable constraint for the identification of phosphopeptides within a set of off-line LC deposited sample spots. It is expected that the development of these methods will not only permit the targeted identification of protein phosphorylation sites but also allow the in-depth analysis of the dynamic events linked to the posttranslational modification.     

Prediction of fluid densities using automatically derived VDW parameters

Automatic parameterization procedures, which are important components in our latest approach of making transferable and extensible general force fields, were demonstrated by deriving parameters from ab initio and empirical data, and subsequently predicting densities for nine molecular fluids. Excellent agreements with available experimental data were obtained for most of the systems studied. The automated procedure for deriving van der Waals parameters, constructed by combining the molecular dynamics simulation and statistical mechanical perturbation theory, is described.     

应用人工神经网络预测多氯代二苯骈呋喃的色谱保留指数

以量子化学方法在密度泛函B3LYP/6-31G(d)水平上计算得到了多氯代二苯骈呋喃系列化合物(PCDF)分子的结构参数:最高占据轨道能(EHOMO)、最低空轨道能(ELUMO)、最正原子净电荷(q+)、最负原子净电荷(q-)、分子偶极矩(μ)、极化率(α)、分子平均体积(Vm)、恒容热容(C■V).采用误差反向传播(BP)算法的人工神经网络,建立了EHOMO、ELUMO、q+、q-、μ、α、Vm、C■V与PCDF色谱保留指数之间关系的模型,检测样本的预报值与实验值相对误差范围为-1.66%².39%,平均相对误差为0.31%,达到了很好的预测效果.     

Mass spectrometry of pancuronium bromide and related quaternary ammonium steroids using the moving belt LC/MS interface

Pancuronium and vecuronium are members of a series of quaternary ammonium steroids used as neuromuscular blocking agents in anesthesiology. In this study the mass spectrometric properties of these bromide salts are examined and spectral features which permit their differentiation are evaluated. The relative merits of chemical ionization and fast atom bombardment using the moving belt liquid chromatography/mass spectrometry interface have been investigated. Fragmentation pathways for both ionization methods were determined with deuterium labeling and linked-scan techniques. Cleaner spectra can be obtained via the matrix-free belt introduction system as compared to conventional fast atom bombardment.     

Prediction of retention of O-ethyl,O-aryl and N-isopropyl phosphoroamidothioates in RP-HPLC from molecular structure parameters

Summary Multi-variable regression analysis between lnk_w, c (in retention equation lnk=lnk_w+cC_b) and molecular structure parameters, including hydrophobicity, electric effect, field effect and position-specific effect constant, was carried out with O-ethyl, O-aryl and N-isopropyl phosphoroamidothioates as test solutes. With these quantitative relationships, the retention behaviour of these solutes for different mobile phase compositions was predicted. The results showed that there are only 26.7% of total, experimentally measured, capacity factors with relative deviations larger than 5% and only 2.2% with deviations larger than 10%, which means that it is possible to apply the method reported to predict retention values for qualitative purposes for different mobile phase compositions.